Abstract 515: Single-cell transcriptomic analysis of HR+/HER2- breast cancer identifies gene signatures that predict outcomes of luminal A and B subtypes

Introduction: Patients with luminal A and B early-stage ER+/HER2- breast cancer (BrCa) are uniformly treated with adjuvant endocrine therapy (ET) (±chemotherapy). A better understanding of drivers of ET resistance is required as subsequent lethal metastatic disease remains a major clinical problem. Therefore, there is an unmet clinical need to identify biomarkers that select low- and high-risk patients who may benefit from de-escalation of current treatments or alternative therapeutic interventions. In the present study, we hypothesized that while luminal A and B tumors both arise from normal hormone-responsive cells, they are transcriptionally distinct, hence allowing the identification of unique gene signatures that can predict outcomes in each subtype. Methods: Tumors from 10 early-stage ER+/HER2- BrCa patients were subjected to single-cell RNA-sequencing (scRNA-seq) analysis (10X Genomics); 6/10 tumors were classified as luminal A and 4/10 as luminal B based on combined PAM50 and immunohistochemical classification (Ki67 cut-off=20%). We performed a direct transcriptional comparison between luminal A and B tumors, using well-established signatures and unbiased differential gene expression analysis. To identify unique luminal A and B tumor-specific genes, we compared the gene expression profile of each luminal subtype with 10 non-neoplastic breast tissues. A predictive model (LASSO) was applied to select genes with the highest frequency using a training dataset. This resulted in 5- and 4-gene signatures for luminal A and B, respectively, which were used to calculate risk scores that divided each subtype into low- and high-risk groups. The prognostic value of the above signatures was validated in an independent dataset. Results: The integrated scRNA-seq analysis of luminal A and B tumors revealed transcriptionally distinct tumor cell clusters while tumor microenvironment (TME) clusters were well intermixed. Luminal B tumors had higher cell cycle and BrCa-specific scores, low ER pathway-gene expression scores, and increased 8q amplifications. IFNγ, OXPHOS, p53, hypoxia and MYC targets were the most upregulated pathways in the luminal B subtype. The TME of luminal B tumors was comprised of lower CD4+ and CD8+ T cell but higher Treg levels. Comparison with normal breast tissues revealed that early-stage ER+ BrCa arises from hormone-responsive epithelial cells and provided a number of tumor-specific genes that were used to generate prognostic signatures. Thes